The steroid binding sites of selected steroid binding proteins will be probed by chemical modification techniques. The proteins to be investigated are the delta5-3-ketosteroid isomerases from Pseudomonas testosteroni and Pseudomonas putida and the progesterone binding protein from rabbit uterine fluids, uteroglobin. Site specific chemical modifications of functional groups will be induced by a variety of techniques including photochemical modifications sensitized by unsaturated ketosteroids which bind to these proteins. The residues modified will be identified in the primary sequence. The effects of these modifications on protein function will be examined in detail, particularly in those instances where covalent attachment of steroid to protein does not occur. The role of carboxyl groups in the catalytic activity of the steroid isomerases will be assessed by specific amidation using a carbodiimide and various amines. In connection with the above studies, the primary structure of P. putida isomerase will be completed. Exploratory studies involving the use of solid phase photoaffinity reagents for labeling steroid binding sites will be conducted.